Genetic disparity between potential hosts and donors limits the utilization and outcome allogeneic BMT by adversely affecting donor availability, engrafument, GVHD, and immune reconstitution. While both non-specific immunosuppression and T cell depletion can decrease frequency and severity of GVHD, no successful program of haploidentical or significantly mismatched unrelated donor BMT has emerged. Further, the non-specific immunologic manipulations and aggressive peri-BMT preparative regimens comprising present methodologies used to ameliorate GVHD are fraught with toxicities including end organ failure, increase in opportunistic infections, B-cell lymphoproliferative disease and loss of graft versus tumor effect. Therefore, attempts to inhibit allorecognitiion specifically while leaving intact the remaining immune repertoire would decrease non-specific toxicity while preserving or improving upon current standards of GVHD control. The central goal of this project is to attempt to selectively inactivate only the small numbers of allospecific T cells transferred in the donor BM that are responsible for GVHD. Alloreactive T cells require two signals for activation. One signal is delivered by alloAg via the TCR and the other by costimulatory molecules. Blockade of B7 family mediated costimulation can induce anergy to fully mismatched allogeneic donor T cells ex vivo. Based on these findings, we have commenced a clinical trial of ex vivo tolerance induction of donor T cells to alloAg. If long-lasting and irreversible unresponsiveness to alloAg can be induced, the associated attendant clinical toxicities of GVHD would be ameliorated while the eligible donor pool would be increased without sacrificing immunity to infectious agents and tumor. To achieve these goals, Four Aims are proposed. First, to continue our ongoing clinical trials of ex vivo anergization of donor T cells to alloAg. Second, block additional pathways to optimize alloAg specific anergy. Third, compare sources of allogeneic T cells to study naive vs previously activated T cells and determine their capacity to be anergized to alloAg. Fourth, determine whether alloanergization results in retention of normal T cell function against pathogens and tumor cells. This Project relies upon and is highly interdependent with the second project (for selection of costimulatory pathways that might block CD4 and CD8 T cell anergy) and with the third project (for preclinical animal models determining costimulatory pathways and study of naive vs previously activated T cells).